Diabetes mellitus is a life-long metabolic disease that can cause several complications representing one of the most important health concerns nowadays. The early diagnosis of Diabetes and regular monitoring of blood glucose level are essential factors in preventing the health complications resulting from this disease. Glycated hemoglobin is an adduct that results from the non-enzymatic reaction of glucose with the N-terminal valine of hemoglobin β-chains. A build-up of glycated hemoglobin within the red blood cell, therefore, reflects the average level of glucose to which the cell has been exposed during its life-cycle and can serve as a marker for average blood glucose levels over the previous months prior to the measurement (J. Y Park et. al. 2008, E. S. Kilpatrick, 2008). In contrast to the common plasma glucose tests, the level of glycated hemoglobin is not influenced by daily fluctuations in the blood glucose concentration but reflect the average glucose levels over the prior six to eight weeks (Goldstein et. al., 2004).
Glycated hemoglobin testing is recommended for checking blood sugar in people who might be pre-diabetic. In fact, the 2010 American Diabetes Association (ADA) added the blood concentration of glycated hemoglobin (HbA1c) of over 6.5% as another criterion for the diagnosis of diabetes (American care 2010). Screening of elevated HbA1c level to a broader population represent an effective way for early diagnosis of diabetes. Higher amounts of HbA1c not only indicate poorer control of blood glucose levels, but also associates with cardiovascular disease, nephropathy, and retinopathy, emphasizing the importance of the precise and accurate monitoring of HbA1C %. Furthermore, monitoring the HbA1c in type-1 diabetic patients may improve treatment (American care 2014).
Current methodologies for HbA1c detection are mainly based on either charge differences (chromatography) (Lafferty J. D. et. al., 2002), structure (affinity or immunoassay assays) or enzymatic assays with the aim to differentiate between HbA1C and native Hb. According to the results of the GH-2 survey of the HbA1c test done by the College of American Pathologists (CAP, USA), immunoassays are the most commonly used methods (65% of participants) followed by the cation-exchange chromatography (31%) then the affinity chromatography (4%). In the chromatography-based methods, the HbA1c % is photometric determined by measuring the ratio of the HbA1c peak area over the tHb peak area. Thus, false positive or negative results can be obtained using these methods due to the possible interferents from the blood (Little R. R. et. al. 2013). Moreover, these methods are generally carried out in centralized laboratories using large and expensive instruments. Several immunoassays have been used for the quantification of HbA1C % using specific mono- or polyclonal antibodies to HbA1C (Wang, B. 2015). Separate ELISA kits based on sandwich assays for the detection of both HbA1c and tHb are commercially available. However, these kits are not intended to produce results for clinical use and cannot be accurately utilised for HbA1C % detection. Immunoassays based on field effect transistor (Qu. L, et. al. 2008) or electrochemical detection using boric acid-modified electrode have been reported (Song S. Y., et. al. 2009). However, the borate-modified electrodes can recognise the blood albumin causing interference. Some sandwich assays using specific HbA1c antibody as capture probe and such lectin or glycan-binding antibodies as detection probe have been reported (Kuno, A. et. al. 2005). However, this method suffers from low sensitivity due the interference from other glycan moieties in the blood sample and the high background signal. In other study, a polyclonal antibody against Hb was used as a common capture probe which binds to all forms of Hb and specific monoclonal antibodies against tHb and GHbA1c were used as detection probes (Chen S. S. et. al. 2012). This immunosensor fabricated using microarray system has eliminated the use of glycan binding molecules and thus, significantly reduced the background interference, achieving high sensitivity. However, immunoassays in general suffers from the instability of the antibodies, their high cost, batch-to-batch variations which limits the clinical usefulness of these methods (Max. V. 2013). Therefore, the development of low cost, stable, portable, specific and simple biosensing platform for the detection of HbA1c is highly demanded and would facilitate the routine monitoring of HbA1c % in blood for the early diagnosis of diabetic patients.
Aptamers are short single stranded DNA (ssDNA) or RNA sequences that have been recently appeared (Ellington A. D. et. al. 1990) as novel recognition receptors which can be used as alternative to antibodies in biosensing devices. Aptamers can be selected in vitro against a variety of targets including small molecules, metal ions and proteins using a process known as SELEX (Sampson, T. 2003). Because of their high affinity and stability, low cost and ease of synthesis with high reproducibility, DNA aptamer are being used as recognition elements replacing antibodies in many biosensing platforms. Recently, Lin H. I. et al. (2015) have reported the first identification of specific aptamers against HbA1c and tHb using microfluidic SELEX chip from a randomized 40-mer DNA library. In this report, the authors have shown a preliminary application of the selected aptamers in an aptamer-antibody sandwich-like chemiluminescence immunoassay. However, the selection of other aptamer sequences for Hb and HbA1C gives a diversity in applying different aptamers that have different molecular structures in a variety of biosensing platforms (McKeague M, et. al. 2014).
Accordingly, there is still a need to develop simple, sensitive, specific, rapid, cost-effective point of care capability of detecting the presence glycated hemoglobin and total hemoglobin in human whole blood.